We have studied human melanoma cell (C8161) adhesion and migration in response to stimulation by soluble collagen IV (CIV) using a modified Boyden chamber. In this modified chamber, shear flow can be introduced over the cell‐substrate interface, affecting tumor cell chemotactic migration through a microporous filter. A relatively high level of intercellular adhesion molecule‐1 (ICAM‐1) was found on C8161 cells. In contrast, levels of β₂‐integrins (e.g., LFA‐1 and Mac‐1), the molecules that would be necessary for C8161 stable adhesion to the endothelium substrate, were found to be very low on these melanoma cells. As a result, C8161 transendothelial migration under a flow condition of 4 dyn/cm² decreased by 70% as compared to static migration. When human neutrophils (PMNs) were present in the tumor cell suspension, C8161 migration recovered by 85% over C8161 cells alone under the 4 dyn/cm² flow condition. Blocking ICAM‐1 on C8161 cells or Mac‐1 on PMNs significantly inhibited C8161‐PMN adhesion and subsequent C8161 migration through the endothelium under flow conditions. In addition, increased interleukin‐8 production and Mac‐1 expression by PMNs were detected when they were co‐cultured with C8161 melanoma cells. These results suggest that transmigration of C8161 cells under flow conditions can be influenced by PMNs, mediated by Mac‐1/ICAM‐1 adhesive interactions and enhanced by altered cytokine production. © 2003 Wiley‐Liss, Inc.